Dendritic cells (DC) exist at the crossroads of innate and adaptive immunity and therefore play a complex and critical role in the host response to infection and in maintenance of self-tolerance. DCs recognize pathogens through a variety of receptors, and this can result in pathogen internalization and rapid production of cytokines that contribute to the inflammatory response. This pathogen recognition also leads to the upregulation of cell surface MHC molecules and costimulatory molecules, which enables DCs to become potent antigen presenting cells presenting pathogen-derived peptides to naive T cells. In this way, DCs are essential for the adaptive immune response, but can also promote autoimmunity if not properly regulated through the inflammatory presentation of self-antigens. DAP12 and FcR3 are two DC-expressed signaling adapters that signal through immunoreceptor tyrosine-based activation motifs (ITAMs). The ITAM-bearing signaling adapters DAP12 and FcR3 regulate DC function in several ways. In vitro, DAP12 and FcR3 inhibit Toll-like receptor (TLR) responses and regulate the anti-inflammatory effects of apoptotic cells on DC function. In vivo, DAP12 and FcR3 regulate DC numbers and contribute to neutrophilia through increased systemic IL-17. Most importantly, DAP12 and FcR3 regulate the maintenance of self-tolerance-DAP12-/-FcR3-/- mice develop lupus-like disease with anti-nuclear antibodies, Ig deposition in the kidney glomeruli and glomerular destruction. The aims of this proposal seek to define the mechanisms by which DAP12 and FcR3 regulate dendritic cell responses in vivo and in vitro with the goal of identifying how these receptors help protect from the development of lupus-like autoimmunity. Specifically, we will 1) identify the FcR3-associated immunoreceptors required for inhibition of TLR responses in conventional and plasmacytoid dendritic cells; 2) test the hypothesis that DAP12 and FcR3 in dendritic cells control neutrophil homeostasis by regulating DC responses to apoptotic cells; and 3) determine if lack of DAP12 and FcR3 specifically in DCs causes the anti-nuclear antibody production and lupus-like disease seen in DAP12-/-FcR3-/- mice. We have shown that DAP12 and FcR3 regulate several aspects of dendritic cell function-TLR signaling, responses to apoptotic cells, and maintenance of self-tolerance-all of which have been associated with the development of lupus-like autoimmunity in mice and humans. The aims of this proposal seek to define the mechanisms by which DAP12 and FcR3 regulate dendritic cell responses in vivo and in vitro with the goal of identifying how these receptors help protect from the development of lupus-like autoimmunity. These studies will not only help elucidate how dendritic cell function is regulated during immune responses, but also how dendritic cell activation is regulated in the steady state. This work will give insights into how to manipulate the immune system to prevent the development of systemic autoimmunity.